Fungal infections continue to be a major cause of morbidity and mortality among HIV-infected patients. Many of the emerging fungal pathogens that are resistant to the three classes of antifungal agents for AID-s-related infections are becoming more common and are associated with higher rates of mortality. Combating drug resistant fungal pathogens should be a top priority. The inhibition of Hsp90 has emerged as a bona fide antifungal target for a number of very important reasons. These include that if increases the efficacy of existing antifungal medications, blocks the emergence of drug resistance, works against a broad spectrum of fungal pathogens, increases survival rates, and pays an important role in attenuating fungal virulence. The development of a Hsp90 fungal inhibitor has been hampered by the fact that fungal and mammalian Hsp90 share structural homology and function. As a result, known Hsp90 inhibitors, such as geldanamycin, induce the heat shock response, an overexpression of Hsp90, Hsp70, and other co-chaperones. This response is both anti-apoptotic and prosurvival, thereby potentially limiting the inhibitors effectiveness as antifungal agents. The challenge is to find inhibitors of Hsp90 that are selective for fungal Hsp90. We have discovered two fungal compounds that are 10X selective inhibitors of Candida Hsp90 relative to mammalian Hsp90. No compounds with this activity have ever been reported. These compounds were screened for using LifePharms' natural product library of 26,000 samples of basidiomycetes and ascomycetes (mushrooms), including nearly every species found in continental United States. The goals of this grant proposal are the following: 1.) to isolate each active compound 2.) to identify and dereplicated lead compounds 3.) to determine the effect of our compounds on stress markers (Hsp90 and Hsp70) in both mammalian and fungal cells 4.) to determine whether our inhibitors interact directly with the ATP binding site on the fungal and mammalian Hsp90 molecule 5.) to determine the in vitro activity and potency of our compounds against panels of clinically important fungal species and 6.) to screen an additional 5,000 extracts for additional fungal-specific Hsp90 inhibitors. The work for goal 5 will be performed by University of Texas Health Science Center at San Antonio. PUBLIC HEALTH RELEVANCE: Fungal infections continue to be a major cause of morbidity and mortality among HIV-infected patients. Antifungal compounds have been discovered from natural products. In collaboration with the Antifungal Susceptibility Testing Laboratory and Division of Infectious Diseases at the University of Texas Health Science Center at San Antonio, we have identified natural products that restore fluconazole toxicity to previously resistant Candida strains.